Polysomes Bypass a 50-Nucleotide Coding Gap Less Efficiently Than Monosomes Due to Attenuation of a 5' mRNA Stem-Loop and Enhanced Drop-off.

Autor: O'Loughlin S; School of Biochemistry, University College Cork, Western Gateway Building, Western Road, Cork, T12 XF62, Ireland; School of Microbiology, University College Cork, Western Gateway Building, Western Road, Cork, T12 YT57, Ireland., Capece MC; Department of Structural Biology, Stanford University School of Medicine, Stanford, CA 94305-4090, USA., Klimova M; Department of Physical Biochemistry, Max Planck Institute for Biophysical Chemistry, 37077 Göttingen, Germany., Wills NM; Department of Human Genetics, University of Utah, Salt Lake City, UT 84112-5330, USA., Coakley A; School of Biochemistry, University College Cork, Western Gateway Building, Western Road, Cork, T12 XF62, Ireland., Samatova E; Department of Physical Biochemistry, Max Planck Institute for Biophysical Chemistry, 37077 Göttingen, Germany., O'Connor PBF; School of Biochemistry, University College Cork, Western Gateway Building, Western Road, Cork, T12 XF62, Ireland., Loughran G; School of Biochemistry, University College Cork, Western Gateway Building, Western Road, Cork, T12 XF62, Ireland., Weissman JS; Department of Cellular and Molecular Pharmacology, University of California, San Francisco, San Francisco, CA 94158, USA., Baranov PV; School of Biochemistry, University College Cork, Western Gateway Building, Western Road, Cork, T12 XF62, Ireland; Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, RAS, Moscow 117997, Russia., Rodnina MV; Department of Physical Biochemistry, Max Planck Institute for Biophysical Chemistry, 37077 Göttingen, Germany., Puglisi JD; Department of Structural Biology, Stanford University School of Medicine, Stanford, CA 94305-4090, USA., Atkins JF; School of Biochemistry, University College Cork, Western Gateway Building, Western Road, Cork, T12 XF62, Ireland; School of Microbiology, University College Cork, Western Gateway Building, Western Road, Cork, T12 YT57, Ireland; Department of Human Genetics, University of Utah, Salt Lake City, UT 84112-5330, USA. Electronic address: j.atkins@ucc.ie.
Jazyk: angličtina
Zdroj: Journal of molecular biology [J Mol Biol] 2020 Jul 24; Vol. 432 (16), pp. 4369-4387. Date of Electronic Publication: 2020 May 23.
DOI: 10.1016/j.jmb.2020.05.010
Abstrakt: Efficient translational bypassing of a 50-nt non-coding gap in a phage T4 topoisomerase subunit gene (gp60) requires several recoding signals. Here we investigate the function of the mRNA stem-loop 5' of the take-off codon, as well as the importance of ribosome loading density on the mRNA for efficient bypassing. We show that polysomes are less efficient at mediating bypassing than monosomes, both in vitro and in vivo, due to their preventing formation of a stem-loop 5' of the take-off codon and allowing greater peptidyl-tRNA drop off. A ribosome profiling analysis of phage T4-infected Escherichia coli yielded protected mRNA fragments within the normal size range derived from ribosomes stalled at the take-off codon. However, ribosomes at this position also yielded some 53-nucleotide fragments, 16 longer. These were due to protection of the nucleotides that form the 5' stem-loop. NMR shows that the 5' stem-loop is highly dynamic. The importance of different nucleotides in the 5' stem-loop is revealed by mutagenesis studies. These data highlight the significance of the 5' stem-loop for the 50-nt bypassing and further enhance appreciation of relevance of the extent of ribosome loading for recoding.
(Copyright © 2020 The Author(s). Published by Elsevier Ltd.. All rights reserved.)
Databáze: MEDLINE
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